Spectrometer and DAD Comparison The apparent discrepancy in the relative extraction efficiency results, 100% for the spectrophotometer and 62% for the DAD, may be explained by the difference in how the data are collected by each instrument. The DAD collects absorbance data at all wavelengths for each separated component as they elute from the column. However, during the data analysis, integration is performed as a function of time for a particular wavelength, ignoring the broad absorbance range characteristic of UV–Vis spectra. Thus, when calculating an absorbance value, the result is not the sum of the absorbance over a range of wavelengths, but for a discrete wavelength as the compound elutes. Additionally, the LC/MS has the ability to separate isomers. Therefore, if a particular dye is a mixture of isomers, the DAD measures their absorbance separately. Conversely, the spectrophotometer collects its absorbance data independent of time. The integration of the spectrophotometer data involves the absorbance over a range of wavelengths and can include isomer contribution. Thus, the data from the DAD are of a single component of the fiber extraction at a particular wavelength, whereas the data from the spectrophotometer are for all absorbing compounds in the selected wavelength band. This is one possible explanation for the difference in the results of the extraction efficiency for the two instruments. A comprehensive evaluation of this difference is beyond the scope of this project. Method Application Basic Method—The optimized LC/MS parameters for fiber dye analysis were tested using the extracts of single fibers. Several fibers of different colors were analyzed in this manner, with their mass spectral and UV–Vis spectra being compared. In some instances, fibers with the same apparent color were shown to have different dye profiles by the resulting chromatograms. In other instances, differences were found only when the mass spectral data were reviewed, thereby demonstrating the additional discriminating power of the LC/MS. The examination of three black acrylic fibers shows the advantage of LC/MS over other methods of fiber dye analysis. The comparison of Fibers A and B demonstrates the superiority of LC/MS over UV–Vis microspectrophotometry in its ability to discriminate between similarly colored items. The UV–Vis spectrum of Fiber A could not be discriminated from Fiber B (see Fig. 3). When the same fibers were extracted and the dyes were analyzed with LC/MS, the isoabsorbance plots illustrated a difference between the two dye profiles (see Fig. 4). For Fiber A, there is an additional absorbance at c. 9.5 min, which is absent from the spectrum of Fiber B. This shows that although Fibers A and B appeared similar from conventional UV–Vis microspectrophotometry, they are, in fact, different.
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